Optimization of adoptive immunotherapy with autologous CD20-specific T cells Non-Hodgkin lymphoma is diagnosed in more than 70,000 Americans each year, and aggressive B-cell lymphomas comprise approximately 35-40% of these cases. Significant proportions of patients relapse or have refractory disease after induction chemotherapy, and the outcomes for these patients are poor. We propose here a novel immunologic therapy for these diseases, using adoptive immunotherapy with genetically modified CD20-specific T cells. CANDIDATE: Dr. Till is an Assistant Member in the Clinical Research Division at the Fred Hutchinson Cancer Research Center (FHCRC) and Assistant Professor in the University of Washington (UW) School of Medicine with 6 years of experience as principal investigator of two phase I adoptive immunotherapy clinical trials using genetically modified T cells. His career goal is to be an independent translational physician-scientist using T cell-based immunotherapeutic to treat lymphoma at a major academic medical center. He gives many indications that he will be successful in his goals, with 6 first-authored peer-reviewed publications, several awards including pilot research grants and career development grants, and a recent appointment to a full faculty position. During the K23 career development period, he intends to address areas of deficiency in laboratory experience, regulatory management, biostatistics, and clinical trial design, using a combination of hands-on experience, formal coursework, intensive workshops, and mentored guidance. The proposed research strategy and 4-year career development plan will provide excellent training for Dr. Till and will allow him to b highly competitive to apply for R01 funding during the last 2 years of the K23 award period. ENVIRONMENT: Dr. Till is fortunate to be in an outstanding position for launching a career in academic medicine. His mentor, Dr. Oliver Press, has >25 years' experience as a leader in translational research developing immunologic therapies for lymphoma, abundant grant funding, and a strong track record of mentoring young investigators to independence. The UW is one of the leading academic medical research institutions in the world, consistently in the top 3 recipients of NIH funding. The FHCRC is an NCI-designated comprehensive cancer center with a focus in immunologic research that has produced 3 Nobel laureates among its faculty. The FHCRC houses world-class research facilities that are available for Dr. Till's use, including a GMP Cell Processing Facility in which clinical grade genetically modified autologous T cells are generated and expanded for use in the clinical trial described in Aim 1, as well as extensive shared equipment and resources encompassing all of the needs for Dr. Till's proposed laboratory experiments. Dr. Till has also assembled an extremely strong team of consultants, including Dr. Stanley Riddell, a world leader in adoptive T cell therapy, Drs. Jeffrey and Martha Ledbetter, experts in T cell biology and development of immunologic therapeutics, and Dr. Ted Gooley, an excellent biostatistician. RESEARCH: The research plan designed by Dr. Till and his mentor is well-suited to developing the skills he needs to become an independent investigator. The clinical trial in Aim 1 will provide valuable experience in navigating regulatory issues such as FDA Investigational New Drug application, NIH Recombinant DNA Advisory Committee, etc., as well as managing the many other facets of a gene therapy trial as principal investigator. In this trial, autologous T cells modified with a lentiviral vector encoding a CD20-specific chimeric antigen receptor (CAR) with CD28 and CD137 (4-1BB) costimulatory domains and an inducible caspase 9 safety switch will be expanded and infused into patients with relapsed or refractory aggressive B cell lymphoma following autologous stem cell transplantation. We believe this approach addresses several of the weaknesses identified in our two previous CD20 CAR clinical trials, and will be a highly effective treatment. Ideally, this adoptive T cell therapy approach would be applicable in the non-transplant setting, and given the difficult nature of treating relapsed or refractory aggressive B cell lymphomas, it is likely that further augmentation of CAR T cell function is needed. In Aims 2 and 3 we outline a novel strategy to improve CAR T cell function through CTLA-4 blockade, in a series of experiments that is designed to provide Dr. Till with intensive experience in the laboratory using a wide array of immunological techniques. In these experiments, T cells will be modified to co-express anti-CTLA-4 scFv-Ig as well as a 3rd-generation fully-human anti-CD20 CAR. The anti-CTLA-4 scFv-Ig will be expressed as either 1) a surface-bound or Golgi-targeted molecule that will effectively prevent or reduce CTLA-4 expression; and 2) as a secreted molecule that hypothesize will have an inhibitory effect on regulatory T cells in the tumor microenvironment. Knockdown of CTLA-4 synthesis using shRNA is another potential strategy to improve the function of CAR-expressing effector T cells through checkpoint inhibition. We are confident that this innovative research plan will provide Dr. Till with a strong transition to independence as an investigator. Furthermore, we are optimistic that the resulting work will have widespread applicability in the field of adoptive T cel immunotherapy, and will ultimately lead to a well- tolerated and potentially curative treatment for aggressive B cell lymphomas.